Improved DNA methylation analysis via enrichment of demethylated cells expressing an X-inactivated transgene

Biotechniques. 2006 Oct;41(4):461-6. doi: 10.2144/000112246.


The demethylating drug 5-aza-2'-deoxycytidine (5-aza-2dC) is frequently used to investigate the effect of global DNA demethylation on gene expression in cultured mammalian cells. Here, we describe a method that uses the reactivation of an X-inactivated green fluorescent protein (GFP) transgene as a marker to enrich for cells that have undergone drug-induced demethylation. By combining it with microarray gene expression profiling, we demonstrate the method's utility in identifying genes activated by global DNA demethylation.

Publication types

  • Technical Report

MeSH terms

  • Animals
  • Animals, Newborn
  • Azacitidine / pharmacology
  • Cells, Cultured
  • DNA Methylation*
  • Female
  • Fibroblasts / drug effects
  • Fibroblasts / metabolism
  • Flow Cytometry
  • Gene Expression Profiling
  • Gene Expression Regulation / drug effects
  • Gene Silencing*
  • Green Fluorescent Proteins / metabolism
  • Mice
  • Oligonucleotide Array Sequence Analysis
  • Organ Culture Techniques
  • Time Factors
  • Transgenes*
  • X Chromosome*


  • Green Fluorescent Proteins
  • Azacitidine